Occurrence of fastidious Campylobacter spp. in fresh meat and poultry using an adapted cultural protocol.

This study used an adapted cultural protocol for the recovery of fastidious species of Campylobacter, to gain a more accurate understanding of the diversity of Campylobacter populations in fresh meats. Chicken (n=185), pork (n=179) and beef (n=186) were collected from supermarkets and butchers throughout the Republic of Ireland. Samples were enriched in Campylobacter enrichment broth for 24h under an atmosphere of 2.5% O(2), 7% H(2), 10% CO(2), and 80.5% N(2). The enriched samples were then filtered onto non-selective Anaerobe Basal Agar supplemented with lysed horse blood using mixed ester filter membranes. Isolates were identified by both genus and species-specific PCR assays and biochemical testing. The incidence of campylobacters on beef (36%) was significantly higher than on pork (22%) or chicken (16%), and far exceeds previously reported prevalence levels. The method was successful in recovering 7 species of Campylobacter, including the fastidious spp. C. concisus and C. mucosalis, from chicken meat, and 10 species, including C. concisus, C. curvus, C. mucosalis, C. sputorum, and C. upsaliensis, from minced beef. The isolation of C. concisus and C. upsaliensis from meat in this study is of particular significance, due to their emerging clinical relevance. The results of this study confirm that the diversity of Campylobacter species on fresh meats is greater than previously reported and highlights the bias of cultural methods towards the recovery of C. jejuni.

Survival characteristics of Cronobacter spp. in model bovine gut and in the environment.

Cronobacter spp. (formally Enterobacter sakazakii) have been linked to illness in infants from contaminated powdered infant formula. The source of the pathogen remains unclear, and it is believed that farm environments and cattle could play a role in the transmission of Cronobacter spp. The aim of this study was to establish if the organism would survive passage through a model of the bovine rumen and abomasum and in bovine feces in the farm environment. Models of the bovine abomasum and rumen were inoculated with Cronobacter strains (final concentrations of 5.7 and 6.5 log(10) CFU/mL, respectively), and survival was examined over time in these environments using an adapted ISO/DTS 22964 culture protocol. Fecal samples were inoculated with Cronobacter (final concentration 6.0 log(10) CFU/mL), and survival on soil and in containers stored outdoors was examined over time. The results showed no significant changes in the number of Cronobacter in rumen fluid over a 24-h period. Cronobacter were undetectable after 30 min of incubation in the model abomasum. The pathogen survived 105 days in sealed containers and was detectable after 112 days in soil. This study indicated that Cronobacter spp. are unlikely to be shed in bovine feces as supported by previous surveillance studies; however, the study did show that the organism survives well in the farm environment.

Surveillance and characterisation by pulsed-field gel electrophoresis of Cronobacter spp. in farming and domestic environments, food production animals and retail foods.

Cronobacter spp. (formally Enterobacter sakazakii) has been linked to illness in infants from contaminated powdered infant formula, however, there is limited information on the environmental sources and potential transmission routes of this pathogen. The aim of this study was to establish if food production animals (cattle, pigs), and the wider farm environment were playing a role in the transmission of Cronobacter spp. and also to assess the risk of cross contamination in the home where infant formula is prepared, from the presence of the pathogen on other foods and the general domestic environment. A wide range of samples (n=518) was collected at dairy farms, meat abattoirs, retail food stores and domestic environs and examined for the pathogen using an adapted ISO/DTS 22964 cultural protocol. The modified method included incubation at 42 degrees C instead of 44 degrees C and serial dilution of the enriched media prior to plating on Druggan-Forsythe-Iversen agar. Presumptive Cronobacter spp. colonies were confirmed by Real Time PCR targeting the dnaG on the MMS operon. All Cronobacter spp. isolated were speciated using biochemical tests, tested for resistance to 8 antibiotics and characterised using pulsed field gel electrophoresis. Cronobacter spp. was not recovered from cattle faeces, farm soil or trough water but isolates (n=33) were recovered from a variety of other sample types including cattle feed, pork and beef cuts, beef burgers and beef mince, green vegetables as well as organic breakfast cereals and domestic vacuum cleaner dust. The species recovered included C. Sakazakii (n=21), C. malonaticus (n=1) and C. turicensis (n=1). Of the 33 isolates 51% were resistant to Cephalothin but sensitive to all other 7 tested antibiotics. Sub-typing of the recovered isolates by PFGE showed considerable clonal diversity, though a number of persistent PFGE profiles were observed. In conclusion the study showed that Cronobacter spp. was not carried by food production animals but was present in a range of diverse sample types and environs with particular association with dry environments.